Anthrax, which is caused by the spore-forming bacterium Bacillus anthracis, has become one of the major bioterrorism threats to our nation. Human vaccination in the USA with licensed protective antigen (PA)-based vaccine, Anthrax Vaccine Adsorbed (AVA), requires six immunizations followed by annual boosters. This underscores the need for development of new, improved anthrax vaccine. The long-term goal of this research is to develop an effective and easily administrated anthrax vaccine, using the B. anthracis protective antigen (PA63), the N-terminal domains of lethal factor (LFn, aminoacids 1-254) and edema factor (EFn, aminoacids 1-254) as vaccine components. Our hypothesis is that an effective vaccine should be composed of multiple relevant antigens delivered by an intranasal route in order to provide mucosal and systemic immunity against anthrax. Formulation of the anthrax vaccine into a nasal spray would allow a mass population to be immunized in a short period at a low cost. Among the currently available mucosal immunization strategies, antigen delivery with a replication-defective adenovirus is a good choice. Recombinant adenovirus and plasmid expression vectors encoding PA63, LFn and EFn will be constructed through this project. In order to evaluate the efficacy of the vaccine and provide an optimal vaccination protocol, intranasal immunization with different combinations of adenoviral vectors will be compared with immunization with plasmid expression vectors by intramuscular injection. The specific aims of this project are: Specific Aim #1: To develop a recombinant adenovirus-vectored multi-component vaccine against anthrax. Specific Aim #2: To compare the systemic and mucosal immunity elicited by the vaccine developed in #1 through intranasal inoculation with that elicited by plasmid expression vectors through intramuscular injection.